Optimization of multi-enzyme cascade process for the biosynthesis of benzylamine

Benzylamine is a valuable intermediate in the synthesis of organic compounds such as curing agents and antifungal drugs. To improve the efficiency of benzylamine biosynthesis, we identified the enzymes involved in the multi-enzyme cascade, regulated the expression strength by using RBS engineering in Escherichia coli, and established a regeneration-recycling system for alanine. This is a cosubstrate, coupled to cascade reactions, which resulted in E. coli RARE-TP and can synthesize benzylamine using phenylalanine as a precursor. By optimizing the supply of cosubstrates alanine and ammonia, the yield of benzylamine produced by whole-cell catalysis was increased by 1.5-fold and 2.7-fold, respectively, and the final concentration reached 6.21 mM. In conclusion, we achieved conversion from l-phenylalanine to benzylamine and increased the yield through enzyme screening, expression regulation, and whole-cell catalytic system optimization. This demonstrated a green and sustainable benzylamine synthesis method, which provides a reference and additional information for benzylamine biosynthesis research.